ABSTRACT Cold Spring Harbor Laboratory proposes to continue the course on X-Ray Methods in Structural Biology to be held in the fall of 2017-2021. X-ray methods, spearheaded by X-ray crystallography, yield a wealth of structural information unobtainable through other methods. This intensive 16-day course combines a thorough theoretical approach with practical experience, including extensive laboratory and computational training. Lectures and practical sessions focus on X-ray crystallographic techniques, which remain indispensable in yielding atomic level structural detail of macromolecules, but also extend to X-ray solution scattering (SAXS), and serial crystallography in X-ray free electron lasers (XFEL) and synchrotron sources. The course aspires to provide the opportunity for researchers working in different biological disciplines to become grounded in the techniques and principles of macromolecular crystallography and complementary methods. Learning the theory and practice of the methods at hand, allows participants with a working knowledge of macromolecular structure and function, but who are new to X-ray methods, to harvest the power of recent developments in automation and apply the methods to their current research questions. Topics in X-ray crystallography include basic diffraction theory, crystallization of soluble and also membrane proteins (including lipidic cubic phase techniques), nucleic acids, macromolecular complexes and viruses, crystal cryo-protection and characterization, home, synchrotron, XFEL X-ray sources and optics, X-ray diffraction data collection and data processing, experimental phasing methods including anomalous diffraction, molecular replacement, density modification, electron density map interpretation and model building, structure refinement and validation, as well as coordinate deposition and structure presentation. Topics in X-ray scattering include theory lectures and computational training to derive SAXS invariants and models to understand macromolecular domain organization. Following three decades that have seen two Nobel prize winners graduating from the course, it is time for change: within the duration of the grant, the organization and running of the course will be handed off to a new generation. Two new instructors will participate in the team from the start, and by the end of the grant period a new set of instructors will ensure continuity and renewal. The course will continue to teach participants through extensive hands-on experiments along with lectures and informal discussions on the theory behind the techniques and the latest developments in automating experimental and computational procedures, from world leaders in methods development. Applications are open to a wide range of students including advanced graduate students, medical students, post-doctoral fellows, faculty at universities and medical schools, as well as Ph.D. and M.D. scientists from industry, aiming to allow students to learn techniques that can be immediately incorporated into their own research, also enabling translational approaches including rational drug design.